Microtubules are essential for a variety of cellular functions, including maintenance of cell shape, cell movement, cell polarity, intracellular transport, mitosis, and meiosis. Microtubule networks exist in dynamic equilibrium and undergo dramatic reorganization through the course of the cell cycle. For example, upon entering mitosis, the interphase microtubule network is rapidly disassembled, followed by the reorganization of microtubules into the mitotic spindle.
Microtubules are comprised of α-β tubulin heterodimers, which are commonly assembled and nucleated by the γ-tubulin protein complexes resided in the centrosomes (Zheng et al., 1995; Erickson and Stoffler, 1996). Regulation of microtubule formation is controlled by two groups of proteins, namely microtubule stabilizers and destabilizers (Heald and Nogales, 2002). The former group is exemplified by microtubule-associated proteins (MAPs), which bind to and stabilize microtubules (Cassimeris and Spittle, 2001). The latter group includes Op18/stathmin (Belmont and Mitchison, 1996; Cassimeris, 2002), katanin (McNally and Vale, 1993; Hartman et al., 1998), and Kin I kinesins (Walczak et al., 1996; Desai et al., 1999), which possess the ability to disassemble microtubules.
Protein 4.1R, originally identified as an 80-kDa protein (i.e., 4.1R-80) in human erythrocytes, plays a crucial role in maintaining the specialized mechanical properties of erythrocyte plasma membranes. Multiple protein 4.1R isoforms, generated by alternative RNA splicing, have been identified in erythroid (Tang et al., 1988; Conboy et al., 1988) and nonerythroid (Tang et al., 1990; Conboy et al., 1991; Huang et al., 1993) cells.
In addition to being associated with the plasma membrane cytoskeleton, protein 4.1R also localizes to intracellular sites in nucleated cells, including the nuclear matrix (De Career et al., 1995; Krauss et al., 1997a), microtubules (Perez-Ferreiro et al., 2001), and centrosomes (Krauss et al., 1997b; Hung et al., 2000). A nonerythroid isoform of 4.1R (4.1R-135) has also been reported to interact with the nuclear mitotic apparatus protein (NuMA) in the interphase nucleus, forming a complex with several spindle-pole organizing proteins, including NuMA, dynein, and dynactin, during cell division (Mattagajasingh et al., 1999). The 4.1R-135 isoform is generated by alternative mRNA splicing, which produces a mRNA encoding a polypeptide having an additional 209 N-terminal amino acid residues, relative to the erythroid 4.1R-80 isoform (Tang et al., 1990). The subcellular localization and the interaction of the nonerythroid 4.1R-135 isoform with NuMA suggest an essential role of 4.1R-135 in organizing the architecture of the nucleus and mitotic spindle (Mattagajasingh et al., 1999).
Using the N-terminal head region of 4.1R-135 (i.e., amino acid residues 1-209) in a yeast two-hybrid screening assay, a centrosomal protein designated centrosomal P4.1-associated protein (CPAP) was identified (Hung et al., 2000). In addition to binding to the head region of 4.1R-135, CPAP is associated with the γ-tubulin complex. Although increasing numbers of factors have been identified that are important for microtubule functions, their mode of action and practical use are yet to be elucidated.